Northeastern Naturalist Vol. 22, No. 1 J. Zaharick, H. Beck, and V. Beauchamp 2015 163 2015 NORTHEASTERN NATURALIST 22(1):163–177 An Experimental Test of Epi- and Endozoochory of Arbuscular Mycorrhizal Fungi Spores by Small Mammals in a Maryland Forest John Zaharick1,*, Harald Beck1, and Vanessa Beauchamp1 Abstract - Small mammals have been proposed as important dispersers of the spores of arbuscular mycorrhizal fungi (AMF), yet few data exist to support this hypothesis. We experimentally tested 2 models of small-mammal dispersal of AMF spores by quantifying their impact on the mycorrhizal inoculum potential of sterile soil flats in a northeast mesophytic forest in Maryland. Epizoochory did not provide a mechanism for spore dispersal in our study. However, our data demonstrated that endozoochory by several small-mammal species can be an effective dispersal mode for AMF. In the field experiment, inoculum potential of soil from plots that were accessible to Peromyscus leucopus (White-footed Mouse) was significantly higher than in control plots, which excluded small mammals. This study provides experimental evidence that White-footed Mice disperse AMF spores. Introduction Mycorrhizal fungi provide nutrients, minerals, and water to plant hosts in exchange for carbon compounds (Leake et al. 2004, Smith and Read 2008). This symbiosis enhances plant productivity and resistance to pathogens (Song et al. 2010, van der Heijden et al. 1998, Vogelsang et al. 2006). Mycorrhizal fungi are known to connect root systems, thus facilitating chemical signaling and transfer of nutrients, hormones, and warnings of predation and disease between individual plants (Song et al. 2010, Whittingham and Read 1982). Individual fungi have been found to connect up to 19 trees simultaneously (Beiler et al. 2010) into networks that may support the growth of juvenile plants in shaded understories (Simard et al. 1997). At the community level, plant diversity can either increase or decrease with the introduction of mycorrhizal fungi (Hartnett and Wilson 1999, Leake et al. 2004) because fungi enhance allelopathy (Barto et al. 2012) and weaken the ability of non-native plants to invade communities (Allen and Allen 1990, Allen et al. 1989). Mycorrhizal-obligate plants, such as members of the family Orchidaceae, depend on the presence of species-specific fungi for germination and survival (McCormick et al. 2004). These influences on the diversity and composition of plant communities depend on the distribution of mycorrhizal fungal species within ecosystems. Mycorrhizal fungi occur in isolated patches around host plants (Peay et al. 2011), and spatial heterogeneity has been shown to influence plant-community composition (Davison et al. 2012, Read 1998). Therefore, the ability of mycorrhizal 1Towson University, Department of Biological Sciences, 8000 York Road, Towson, MD 21252. *Corresponding author - johnzaharick@gmail.com. Manuscript Editor: Kurt Moseley Northeastern Naturalist 164 J. Zaharick, H. Beck, and V. Beauchamp 2015 Vol. 22, No. 1 fungi to disperse may govern their distribution and ultimately, the distribution and ecology of plants. Epigeous fruiting bodies, such as mushrooms and puffballs, can directly release spores into the air, but fungi with hypogeous fruiting bodies have no obvious means of dispersing spores (Johnson 1996). Small mammals have been considered important for dispersing mycorrhizal fungal spores (Cázares and Trappe 1994, Frank et al. 2006, Vernes and Dunn 2009) because they consume hypogeous fruiting-bodies (Mangan and Adler 2000, Vernes and Dunn 2009). Spores remain viable and can inoculate greenhouse plants after passing through mammalian digestive systems (Colgan and Claridge 2002, Johnson 1996, Reddell et al. 1997, Trappe and Maser 1976). However, few experimental studies have tested the extent to which small mammals disperse mycorrhizal fungi in the field. For example, in an experiment at Mt. St. Helens, WA, aerial-spore traps only collected arbuscular mycorrhizal fungi (AMF) spores near Thomomys talpoides Richardson (Pocket Gopher) mounds (Allen 1987). When Pocket Gophers were placed in enclosures, plants within the fencing became inoculated with AMF whereas plants immediately outside the enclosures remained AMF free (Allen and MacMahon 1988). However, the dispersal mechanism may have been Pocket Gophers bringing buried spores to the surface where spores were then dispersed by the wind (Allen 1987). Animal effects on AMF communities were found in an Australian rainforest where small-mammal exclosures had lower mycorrhizal inoculum potential, spore abundance, and spore species richness after a 3-year period than accessible controls (Gehring et al. 2002). It remains unclear though whether small mammals were directly dispersing spores or indirectly increasing mycorrhizal levels by killing seedlings and severing roots because dead roots may act as a greater source of inocula than living roots (Gehring et al. 2002). Because the results of these studies are inconclusive, further research is necessary to test whether the spores of mycorrhizal fungi are dispersed by small mammals. If small mammals are the primary vector of mycorrhizal fungi, then they may play a critical ecological role in determining the dis tribution, fitness, and species richness of mycorrhizal plants (Gehring et al. 2002, Leake et al. 2004), as well as soil-nutrient levels and soil-fauna abundance (Coleman et al. 2004). Small mammals may also be necessary for dispersing mycorrhiza-forming fungal spores to areas where mycorrhizal activity has been reduced after disturbance, such as by windthrows, landslides, fires, floods, herbivore browsing, or human activities (Boerner et al. 1996, Bressette et al. 2012, Fisher and Fulé 2004, Perry et al. 1987, Wearn and Gange 2007). In this study, we conducted a mark–recapture survey and a field experiment to determine the extent to which epizoochory and endozoochory contribute to AMF spore dispersal by small mammals in a northeastern mesophytic forest. We hypothesized that the inoculum potential of soil in plots accessible to small mammals would be higher than control plots where small mammals were excluded. Northeastern Naturalist Vol. 22, No. 1 J. Zaharick, H. Beck, and V. Beauchamp 2015 165 Field-site Description The study site was located at the Towson University (TU) Field Station in Monkton, MD (39°35'57"N, 76°37'40"W; Fig. 1), which was established in 2010 and comprises 92 ha of closed-canopy mesophytic forest. The station is located in the Piedmont Plateau Province of northern Baltimore County at an elevation of Figure 1. Topographic map of the Towson University Field Station in Maryland (Roberge 2011). Grids A, B, and C were trapped in April, June, August, and October 2011. Only grid B was trapped in 2012. Circles 1–10 are locations for the expe rimental plots. Northeastern Naturalist 166 J. Zaharick, H. Beck, and V. Beauchamp 2015 Vol. 22, No. 1 approximately 136 m. The station contains steep, stream-carved ravines with silty loam soils. The dominant tree species includes Liriodendron tulipifera L. (Tulip Poplar), Fagus grandifolia Ehrh. (American Beech), and Acer rubrum L. (Red Maple). The forest understory is sparse; the most common species are: Chimaphila maculata Pursh (Spotted Wintergreen), Mitchella repens L. (Partridgeberry), and Polystichum acrostichoides (Michx.) Schott (Christmas Fern). To the northwest and southeast, the field station connects to a riparian forest that forms the 7200-ha Gunpowder Falls State Park. Methods Small-mammal trapping We conducted small-mammal live-trapping in April, June, August, and October 2011 and 2012. We established 3 trapping grids ~150 m apart along a southwest–northeast transect in the central portion of the TU Field Station (Fig. 1). Each grid consisted of 100 Sherman traps (H.B. Sherman Traps, Inc., Tallahassee, FL) in a 10 x 10 arrangement with 10 m between traps. Grids contained 50 large traps (8.9 cm x 7.6 cm x 22.9 cm) and 50 small traps (5.1 cm x 6.3 cm x 16.5 cm) placed in an alternating pattern. Bait used in traps contained equal parts peanut butter, paraffin wax, rolled oats, and raisins. We added polyester stuffing to traps in April and October to serve as nesting material when night-time temperatures approached freezing. We placed and monitored 1 grid of traps at a time and checked them for 4 consecutive days beginning at sunrise. We re-baited and reset traps as needed, and later moved them to the next grid. To avoid potential sampling bias, we shifted the order in which we surveyed grids each month. Because the species-accumulation curve reached an asymptote at the end of 2011, we surveyed only 1 grid in 2012. We identified captured individuals to species and attached a uniquely numbered 1005-1 monel ear-tag (National Band and Tag Company, Newport, KY) to each animal before release. We recorded the trap location of all captures and the ear-tag number of previously marked individuals. The handling protocol followed the Society of Mammalogists’ guidelines for using mammals in research (Sikes et al. 2011). To determine the rate and magnitude of epi- and endozoochory, we collected 1 fur and 1 fecal sample per captured individual once per month. We collected fur samples using a piece of transparent adhesive tape (1.5 cm x 6.5 cm) placed against each individual, covering both the ventral and dorsal surfaces. We then affixed the tape samples to a microscope slide in the field and examined it at 100x magnification with a compound light microscope for the presence of AMF spores. We collected fecal samples when animals defecated during handling and stored them at 4 ºC at Towson University prior to processing. To extract spores from fecal samples, we ground 2–3 fecal pellets from a single individual and stirred the ground material in a glass vial containing 75% ethanol. We filtered the resulting mixture twice using 250-μm and 45-μm USA standard test-sieves (Newark Wire Cloth Company, Clifton, NJ). Material from the 45-μm sieve was rinsed into a Petri dish and examined at 20x magnification using a dissecting microscope. We transferred Northeastern Naturalist Vol. 22, No. 1 J. Zaharick, H. Beck, and V. Beauchamp 2015 167 spores from the dish via pipette to a microscope slide containing a drop of Melzer’s reagent for preservation and identification (Maser et al. 1978). Aerial-spore sampling To quantify the abundance and seasonal variation of AMF spores dispersed by wind, we placed eight 15 cm x 1.5 cm Petri dishes containing adhesive paper in each trapping grid in April, June, August, September, and October 2011. We divided each grid into 4 segments and randomly placed 2 dishes within each segment for a 48-hour period (Allen 1987). Dishes were laid flat on the forest floor. After exposing Petri dishes to the air at the field station, we examined them at 20x magnification and transferred AMF spores to microscope slides containing Melzer’s reagent for preservation and identification. Experimental procedure Data from the 2011 field survey indicated that AMF spores occurred in smallmammal feces during June and August, but not in April or October. Therefore, we carried out the experiment from May through September 2012. We established 10 plots by placing 10 parallel lines over a map of the field station, with ~50 m between lines (Fig. 1). We randomly placed 1 plot on each line; each plot contained three 1-m2 subplots with 10 m between subplots. Small mammals had access to 2 experimental subplots and were excluded from 1 control subplot. To encourage small-mammal activity, 1 open subplot contained bait of the same mixture used in trapping, and the other open subplot was not baited. We placed bait in sub-plots once per week from May to the end of September 2012. Small-mammal exclosures consisted of aluminum flashing extending approximately 60 cm aboveground and 10 cm below-ground to prevent mammals from entering. We placed two pieces of rebar in each corner of the subplot to support the flashing—one inside the structure, one outside, and both tied together with wire— and stapled 18 x 16-mesh window screen over the tops of exclosures to prevent mammals from entering. Each subplot contained a single 56 cm x 28 cm x 5.5 cm plastic tray filled with soil sterilized in an electric soil sterilizer by heating it to 82 °C for 2 hours (Kawamoto and Habte 2011). We placed the plastic trays in shallow depressions so their tops were at ground level. We placed metal hexagonal fencing over the soil trays and held it down with metal stakes to prevent large mammals from digging in subplots and displacing soil. We placed a 20-cm-long, 10-cm-diameter, longitudinally cut piece of green PVC pipe in each subplot to serve as shelter for animals because small mammals typically avoid open spaces where they are vulnerable to predators (Catall et al. 2011, Perea et al. 2011). We autoclaved pipe sections at 121 °C for 30 minutes to avoid contamination of the experimental soil. All sites occurred on sloped surfaces where AMF spores present in the soil could be dispersed by rainwater flowing downhill. Therefore, we placed u-shaped plastic rain-guards 50 cm uphill of accessible subplots to redirect surface flows of rainwater away from subplots. Northeastern Naturalist 168 J. Zaharick, H. Beck, and V. Beauchamp 2015 Vol. 22, No. 1 To monitor aerial-spore deposition near subplots and assess its contribution to inoculum potential, we placed 1 aerial-spore trap within 1 m of each subplot for a 7-day period each month for the 5 months of the experiment. We checked exclosures once a week for damage and conducted repairs as needed. To document whether small mammals interacted with soil trays, we deployed motion-activated cameras (Moultrie Game Spy D-40, Alabaster, AL) on baited subplots for a 1-week period each month from May to September 2012. We mounted cameras ~45 cm above the ground on either wooden stakes or rebar, and placed them 50 cm away from baited subplots. We angled the cameras so that they could photograph the entire subplot. We took a 1-L soil sample from the surface of each of the 30 subplots to collect feces during the first week of October 2012 and transferred it to the Towson University greenhouse. We followed Anderson et al. (2010) to measure levels of mycorrhizal fungi in the soil and detect mycorrhizal inoculum potential by growing Sorghum bicolor drummondii (Nees ex Steud.) de Wet and Harlan (Sudangrass) in soil samples placed in bleach-disinfected Deepots. We harvested plants 30 days after seed germination, which provided sufficient time for primary AMF root colonization while limiting secondary AMF colonization (INVAM 2013a, Moorman and Reeves 1979). We removed fine roots and fixed them in 75% ethanol (Beauchamp et al. 2006). Roots were cleared in 5% KOH, stained in Trypan blue (Koske and Gemma 1989), and placed on microscope slides containing polyvinyl alcohol lactoglycerol mounting medium (INVAM 2013b). We measured root-length colonization by mycorrhizal fungi using the grid-line-intersect method (Giovannetti and Mosse 1980). Statistical analysis We used a null-hypothesis statistical testing approach with significance set to P < 0.05 to examine differences between treatments. Treatment type and plot number were the independent variables, and presence of fungi was the dependent variable. We ran a model to explain mycorrhizal inoculum potential using a variable- dispersion beta distribution as recommended for proportion values bounded between 0 and 1 (Cribari-Neto and Zeileis 2010), with a complementary log-link function, and increased zero values by 0.0001 to conform to this distribution. We examined heteroskedasticity of data with a simple linear regression model and a studentized Breusch and Pagan test. The models were run in software program R (R Development Core Team 2010). Results Evidence of epizoochory Adhesive tape used to investigate epizoochory picked up mammal fur and ectoparasites, but out of 223 samples, we detected only a single AMF spore on a Peromyscus leucopus Rafinesque (White-footed Mouse) in June 2012. Northeastern Naturalist Vol. 22, No. 1 J. Zaharick, H. Beck, and V. Beauchamp 2015 169 Evidence of endozoochory We found AMF spores in the feces of White-footed Mice, Sciurus carolinensis Gmelin (Eastern Gray Squirrel), and Microtus pinetorum LeConte (Woodland Vole). We dectected a total of 353 AMF spores in 71 samples of small-mammal feces collected from June and August 2011. Spore frequency varied in samples containing spores from 9 fecal samples containing only a single spore each to 2 samples containing over 100 spores each. We detected a total of 12 AMF spores in 57 samples collected from June and October 2012. Feces of White-footed Mice and Woodland Voles contained the majority of spores detected (Table 1). One Eastern Gray Squirrel sample contained 1 spore in June 2011. We identified all AMF spores to the genus Glomus (Schüßler and Walker 2010) and further classified them into 8 morpho-species based on color, spore size, and spore-wall thickness. Two spores were too degraded to place in a morpho-species. One morpho-species was only found in August and September, and another was only found in a Woodland Vole. Wind dispersal AMF spores dispersed via wind at a rate of 0.77 spores/m2/day in 2011 and 0.74 spores/m2/day in 2012. The 120 aerial-spore traps placed in grids in 2011 collected a total of 4 spores in April and September. The 150 aerial traps placed near experimental subplots in 2012 collected 15 spores, 7 of which were in August (Table 2). At 4 of the subplots where we detected wind-dispersing spores, AMF also appeared in the bioassay. Six plots where wind-dispersed fungi were detected contained no AMF in the bioassay. We detected no wind-dispersed spores in 7 subplots with AMF in the soil (Table 3). Table 2. Number of arbuscular mycorrhizal fungal (AMF) spores detected in aerial spore traps per month in a Maryland mesophytic forest from April to October 2011 (n = 24) and May to September 2012 (n = 30). Year/month AMF spore counts Year/month AMF spore counts 2011 2012 April 2 May 2 June 0 June 1 August 0 July 4 September 2 August 7 October 0 September 1 Table 1. Occurrence of arbuscular mycorrhizal fungal (AMF) spores in White-footed Mouse and Woodland Vole fecal samples (n = 128) in a Maryland mesophytic forest in June and August 2011 and June and October 2012. June 2011 August 2011 June 2012 October 2012 # of samples (% containing AMF) 35 (14.3%) 36 (16.7%) 29 (6.9%) 28 (3.6%) Median abundance of spores 1 1 3 6 Range of abundance of spores 1–134 1–192 3 6 Northeastern Naturalist 170 J. Zaharick, H. Beck, and V. Beauchamp 2015 Vol. 22, No. 1 Experimental results Camera traps aimed at baited subplots documented White-footed Mice, Eastern Gray Squirrels, and Procyon lotor L. (Raccoon) at all baited subplots. Tamias striatus L. (Eastern Chipmunk), Didelphis virginiana Kerr (Virginia Opossum), Marmota monax L. (Groundhog), and Felis catus L. (Feral Cat) also appeared at some subplots. In 4 subplots, roots from the surrounding soil grew into soil trays from below through drainage slots. We detected AMF in 2 subplots in which foreign roots entered the sterilized soil. Unknown animals broke into 4 exclosures and disturbed the sterilized soil. One of those exclosures contained AMF and the other 3 did not. Soil from experimental subplots resulted in more AMF infections than soil from controls (4 baited, 5 non-baited, 2 control). A studentized Breusch and Pagan test showed that a simple linear regression model of our data exhibited heteroskedasticity (BP11 = 19.341, P = 0.05). The variable-dispersion beta regression model was therefore used because it is naturally heteroskedastic. The heteroskedasticity in our data was most likely due to the large number of 0 observations in treatments (Fig. 2). Therefore, following Cribari-Neto and Zeileis (2010), we ran the model with treatment as an additional regressor: percent infected = treatment + plot | treatment This model was significant for both the baited (Z15 = 4.456, P < 0.001) and nonbaited (Z15 = 4.577, P < 0.001) accessible plots. Table 3. Number of arbuscular mycorrhizal fungal (AMF) spores detected in aerial-spore traps placed at experimental subplots, and AMF infections detected in bioassay per subplot in a Maryland mesophytic forest from May to September 2012 (n = 30). Subplots that had neither aerial-dispersing spores nor soil fungi are not listed. A was the exclosure, B the baited accessible subplot, and C the non-baited accessible subplot. Subplot AMF aerial spore counts Fungi in bioassay (%) 7A 1 0.30 9A 0 0.96 4B 0 0.41 6B 4 9.75 7B 2 0.00 8B 1 0.00 9B 0 3.69 10B 0 2.32 2C 1 0.00 3C 2 0.00 4C 0 4.09 5C 1 0.00 6C 1 0.31 7C 1 0.32 8C 0 14.51 9C 0 0.29 10C 1 0.00 Northeastern Naturalist Vol. 22, No. 1 J. Zaharick, H. Beck, and V. Beauchamp 2015 171 Discussion Small mammals play key roles in ecosystems via herbivory, preying on and dispersing seeds, and as a food source for carnivorous species (Kaminski et al. 2007). It is widely assumed that small mammals also disperse mycorrhizal fungi spores and therefore are critical in influencing successional processes and plant-community structure (Cázares and Trappe 1994, Frank et al. 2006, Janos et al. 1995, Vernes and Dunn 2009). Our results provide experimental support of endozoochorial dispersal of AMF by White-footed Mice in northeastern mesophytic forests. We found AMF spores in 3.6–16.7% of fecal samples from White-footed Mice and Woodland Voles (twice in amounts of over 100 spores per sample), and inoculum potential of soil in plots accessible to small mammals was higher than in plots where these mammals were excluded. Presence of significant spore quantities in White-footed Mouse and Woodland Vole fecal samples demonstrates the ability of small mammals to disperse AMF spores in a more concentrated fashion than wind, increasing the likelihood of viable Figure 2. Plot of residuals from the linear model percent infected = treatment + plot versus percent of mycorrhizal fungi infection observed in experimental treatments. Due to the low detection rate of fungi, most observations were 0, which made the data heteroskedastic. Northeastern Naturalist 172 J. Zaharick, H. Beck, and V. Beauchamp 2015 Vol. 22, No. 1 spores being placed near plants (Maser et al. 1978). Such concentrated fecal–spore mass also makes mammalian spore dispersal patchy, as opposed to a more homogenous, but potentially unidirectional, distribution in wind dispersal. This dispersal mechanism has the potential to affect plant succession by adding heterogeneity to the landscape. In an Estonian temperate forest, AMF richness and community composition varied spatially in plots 30 m away from each other, and the overlying plant community reflected this (Davison et al. 2012). In addition, wind-dispersed spores might not necessarily reach all parts of a forest due to prevailing wind direction. Small mammals could be vital in dispersing spores to forest areas where wind dispersal of spores would not likely occur. Our results also suggest that small-mammal dispersal of AMF spores primarily occurs through endozoochory. We found only one Glomus spp. spore among 223 fur samples, suggesting that AMF spores are not dispersed via the fur of White-footed Mice and Woodland Voles. Although epizoochory has been studied in invertebrates (Lilleskov and Bruns 2005, Warner et al. 1987), ours was the first study to test epizoochory as a dispersal mechanism for AMF in small mammals. We encountered White-footed Mice at every baited subplot in the field experiment. Wind-dispersed spores may have inoculated some subplots, but fungi did not appear in the experimental soil in 6 subplots where spores were detected in the air. A third potential source of AMF in the field experiment included fungi present in plant roots that entered experimental trays. We took the top layer of soil from every subplot because that is where we expected to find fecal matter. The influence of foreign roots growing from below was likely minimal because we did not collect most of the soil they had contacted. We could not make direct comparisons between mammalian and wind-mediated dispersal rates because we did not measure the rate of defecation. Janos et al. (1995) determined that Proechimys spp. (spiny rats) defecated 1.3 g dried feces/day/100 g body mass in a Peruvian Amazon rainforest; however, defecation rates for Whitefooted Mice are unknown. While spiny rats and Oryzomys spp. (rice rats) were calculated to disperse 7.30 x107 Glomus spores/ha/yr, Janos et al. (1995) found AMF in 69.3% of all fecal samples. We observed AMF in only 6.6% of fecal samples, suggesting a much lower dispersal rate than what may occur in the Peruvian rainforest. To compare dispersal rates indirectly, Glomus spp. spores precipitated from the air at a consistent rate both years. In contrast, mycophagy varied between years, with the proportion of small mammals detected consuming fungi lower in 2012 than 2011. Detection of AMF spores in small mammals and the wind roughly peaked in the summer months with fewer spores in spring and autumn. This pattern coincides with the fruiting phenology of host plants, when AMF are known to maximize spore production (López-Sánchez and Honrubia 1992). Studies on AMF phenology are needed to better understand seasonal and annual variation in spore production. We may have detected fewer small mammals consuming AMF in 2012 for reasons other than a change in foraging behavior. Mycorrhizal fungi can compose as little as 1% of White-footed Mouse and vole diets by volume (Whitaker 1962). The small mammals examined in this study may have unintentionally consumed Northeastern Naturalist Vol. 22, No. 1 J. Zaharick, H. Beck, and V. Beauchamp 2015 173 spores while feeding on other material as opposed to species such as Glaucomys sabrinus Shaw (Northern Flying Squirrel) or Clethrionomys Tilesius (Red-backed Vole), which consume mycorrhizal fungi as a large portion of their diet (Maser et al. 1978, Smith 2007). Mycophagy may have occurred and not been detected as the concentration of of spores in Peromyscus maniculatus Wagner (Deer Mouse) feces peaks less than 12 h after animals consume fungi and reaches half concentration in another 12 h (Cork and Kenagy 1989); thus, our tests on the feces of animals that had not consumed fungi within a day of capture would likely produce negative results. All spore counts in this study represented minimum counts for what small mammals are able to disperse. We used fewer traps in 2012, resulting in fewer fecal samples, which reduced the likelihood of detecting mycophagy. We were also unable to trap large numbers of species other than White-footed Mice. We trapped only 4 Woodland Voles in 2 years. Alternate trapping methods, such as pitfall traps, may be a better method for sampling this species (Wilson et al. 1996). Small mammals may be among many vectors of AMF, each dispersing a small amount of inocula, but in sum, dispersing large amounts of fungal reproductive structures across different distances and areas (Fig. 3). Collembola (Springtails) disperse AMF-hyphae fragments (Klironomos and Moutoglis 1999, Seres et al. 2007), Formicidae (ants) concentrate root fragments containing AMF in their nests (Friese and Allen 1993, Harinikumar and Bagyaraj 1994, McIlveen and Cole 1976), and in one study, Leporidae (rabbit) feces and whole individuals of Orthoptera (grasshoppers) inoculated plants with AMF (Ponder 1980). AMF spores have also been found in mud nests of Turdus migratorius L. (American Robin), Hirundo Figure 3. Conceptual model of distances over which biotic and abiotic vectors can transport arbuscular mycorrhizal fungi spores and hyphae. Animals may consume fungi (mycophagy) or transport soil containing fungi when constructing nests. Spores brought to the surface may then be dispersed by wind; alternatively, water may directly remove spores from the soil in runoff, especially during floods (see citations in text for distances ). Northeastern Naturalist 174 J. Zaharick, H. Beck, and V. Beauchamp 2015 Vol. 22, No. 1 erythrogaster L. (Barn Swallow), and Trypoxyloninae (mud-dauber wasps) (McIlveen and Cole 1976). Johnson (1996) suggested that spores are wind-dispersed after being exhumed by small mammals. Ants have also been cited in this capacity (Harinikumar and Bagyaraj 1994, McIlveen and Cole 1976), and wind has been found to disperse spores up to 2 km in open habitats (Allen et al. 1989, Warner et al. 1987). Spores and hyphae can also be transported in flood deposits, dispersing potentially up to 100 km (Harner et al. 2009). However, such dispersal would be restricted to riparian areas. Previous research has shown that small-mammal feces can inoculate vascular plants with AMF in greenhouse conditions (Reddell et al. 1997, Trappe and Maser 1976) and small mammals are able to increase AMF-spore species richness and inoculum potential in the field (Gehring et al. 2002). Our data demonstrate that sterile soil exposed to small mammals in a mesophytic forest habitat became inoculated with AMF spores, and those spores were capable of establishing a mutualistic relationship with plants. Small mammals may serve as important vectors for dispersal of mycorrhizal fungi and potentially play a critical role in the distribution, fitness, and species richness of mycorrhizal plants. Acknowledgments We thank A. Henneman for permission to conduct research on his property and D. Forester for assistance in working at the Towson University Field Station. J. Snodgrass provided input on the experimental design. Thanks to A. Cannavino, D. Engel, K. Gazzara, C. Graff, R. Hebert, M. Kopansky, B. Link, A. Marcangeli, K. Michael, L. Motier, J. Peterson, A. Simon, B. Summers, and N. Wentz for work in the field. 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